The major goal of this study is to identify the molecules involved in the intercellular adhesion of teratocarcinoma stem cells and to determine the role these molecules may play during the differentiation of teratocarcinoma stem cells. We have previously identified a fucan/mannan-specific lectin present on the stem cell surface with a proposed role in stem cell intercellular adhesion. As previously described, we have identified the lectin as a protein of 56 kilodaltons. Studies for the next year will focus upon localizing the lectin during teratocarcinoma differentiation in vitro and during mouse pre-implantation development and substantiating its role in the intercellular adhesion of teratocarcinoma stem cells. Toward this goal, we are in the process of characterizing a polyclonal antibody raised against the 56 kilodalton protein. This antibody will then be used in immunocytochemical studies (immunofluorescence of whole cells and of frozen sections) to determine if the lectin is present on the surface of stem cells, the differentiative derivatives, endoderm, and the various cell types present in the pre-implantation mouse embryo. Biochemical studies using immunoprecipitation of metabolically-labeled material followed by one-\and two-dimensional gel analysis will examine the expression of this protein during differentiation. Cell type specificity may also be obtained without employing the antibody by identifying the lectin on two-dimensional gels and then determining if this protein is expressed by the various cell types. To evaluate the role of the lectin in stem cell adhesion, Fab fragments of the antibody will be used to attempt to block the adhesion of stem cells reaggregating in rotary culture. We are also interested in determining the role the lectin may play during stem cell differentiation. Preliminary studies suggest that carbohydrate antagonists of the lectin (fucans) specifically inhibit endoderm differentiation without effecting cell division. Further studies will examine the specificity of this effect more closely and evaluate the extent of inhibition. In addition, we propose to determine the effect of the lectin-specific antibody (whole antibody and Fab fragments) upon stem cell differentiation. (A)